Preparation of A Magnetic Nanosensor Based on Cobalt Ferrite Nanoparticles for The Electrochemical Determination of Methyldopa in The Presence of Uric Acid.

AIM AND OBJECTIVE: Methyldopa is one of the medications that is used for the treatment of hypertension. Therefore, the determination of methyldopa in the presence of other biological components is essential. In this work, a promising electrochemical sensor based on CoFe2O4 magnetic nanoparticles modified glassy carbon electrode (CoFe2O4/GCE) was developed for electrochemical determination of methyldopa in the presence of uric acid. Cobalt ferrite nanoparticles were synthesized via chemical method. MATERIALS AND METHODS: Characterizing the CoFe2O4 was investigated by field emission scanning electron microscopy (FESEM), X-ray diffraction (XRD), energy dispersive X-ray spectroscopy (EDX), transmission electron microscope (TEM), and cyclic voltammetry techniques. RESULTS: Under the optimal experimental conditions, the current response of the electrochemical sensor obtained with differential pulse voltammetry was increased linearly in the concentration range from 1.45 to 15.1 µmol L-1 with the detection limit of 1.07 µmol L-1 for methyldopa. Also, by using the proposed method, methyldopa and uric acid could be analyzed in a mixture independently. The difference in peak potential for analytes is about 150 mV. CONCLUSION: The present sensor was successfully applied for the determination of methyldopa in the presence of uric acid in biological samples and the pharmaceutical samples with satisfactory results.

First report for voltammetric determination of methyldopa in the presence of folic acid and glycine.

In this study, a carbon paste electrode modified with TiO2 nanoparticles and ferrocene monocarboxylic acid (FM) was used to prepare a novel electrochemical sensor. The objective of this novel electrode modification was to seek new electrochemical performances for the detection of methyldopa in the presence of folic acid and glycine. The peak potentials recorded in a phosphate buffer solution (PBS) of pH7.0 were 325, 750 and 880 mV vs. Ag/AgCl/KCl (3.0M) for methyldopa, folic acid and glycine, respectively. Under the optimum pH of 7.0, the oxidation of methyldopa occurred at a potential about 160 mV less positive than that of the unmodified carbon paste electrode (CPE). The response of catalytic current with methyldopa concentration showed a linear relation in the range from 2.0×10(-7) to 1.0×10(-4)M with a detection limit of 8.0 (± 0.2)×10(-8)M.

High sensitive and selective HPTLC method assisted by digital image processing for simultaneous determination of catecholamines and related drugs.

A highly sensitive and selective thin layer chromatographic (TLC) method was developed for simultaneous determination of catecholamines and their related drugs using a new detection method and digital image processing of chromatographic plates. For the quantitative evaluation of the investigated compounds, the chromatographic separation was followed by spraying the plate with 0.02% solution of 2,2-diphenyl-1-picrylhydrazyl radical (DPPH) in ethanol. The BioDit Thin Layer Chromatography (TLC) Scanner device and advanced specific software (ImageDecipher-TLC, Sorbfil TLC Videodensitometer and JustTLC) were used for the detection and quantification of chromatographic spots. For an accurate determination, the RGB colored images of the bright-white spots detected against a purple background were inverted and processed after their conversion into green scale. The results showed a strongly linear correlation between area (R(2)>0.99) and volume (R(2)>0.99) of spots and concentration of investigated compounds in all cases. The limit of detection (LOD) and the limit of quantification (LOQ) were below 49.3 ng/spot and 69.6 ng/spot respectively in all cases. The evaluation of the method was performed using different pharmaceutical samples spiked with the investigated amines and validated with respect to accuracy and precision.

Adsorptive stripping voltammetry determination of methyldopa on the surface of a carboxylated multiwall carbon nanotubes modified glassy carbon electrode in biological and pharmaceutical samples.

In the present work, a simple carboxylated multiwall carbon nanotubes (CMWCNTs) modified glassy carbon electrode was developed for sensitive determination of methyldopa (MTD). The study of modified electrode and MTD electrochemical behavior at its surface was investigated employing SEM, adsorptive stripping voltammetry, electrochemical impedance spectroscopy and chronocoulometry. These studies show that the oxidation of MTD is facilitated at the surface of GCE which is casted with CMWCNTs and remarkably peak current enhanced comparing to the bare electrode due to its adsorption on the electrode surface. Also, because of the catalytic property of modified electrode onset potential decreased for oxidation of MTD. Under optimized conditions, the calibration curve was linear in two concentration ranges of 0.1-30 and 30.0-300.0 µM with a detection limit of 0.08 µM and relative standard deviation (R.S.D.%) lower than 3.0% (n=5). This modified electrode was used as a sensor for determination of MTD in pharmaceutical and human urine samples with satisfactory results.

A peroxidase-based method for the determination of dopamine, adrenaline, and α-methyldopa in the presence of thyroid hormones in pharmaceutical forms.

The analytical properties of two commercial plant peroxidases isolated from horseradish roots and soybean hulls in the catalysis of the transformation of some catecholamines were demonstrated in the absence and presence of thyroid hormones (l-thyroxine and 3,3',5'-iodothyronine). For the first time the reactions of dopamine, adrenaline, and α-methyldopa oxidation with H(2)O(2) catalysed by horseradish peroxidase with the addition of l-thyroxine as the amplification agent were studied and proposed as the indicator reactions for the simple and rapid enzymatic determination of the indicated catecholamines in their concentration ranges 0.5-300, 4-300, and 100-400 µM, respectively. The catalytic activity of the enzyme (characterized by the reaction rate) was controlled spectrophotometrically. The optimum conditions for the indicator reactions were thoroughly characterized. The mechanism of the stimulatory effect of l-thyroxine on the oxidation of the catecholamines was discussed. The developed enzymatic procedures were successfully applied for the determination of dopamine, adrenaline, and α-methyldopa in some pharmaceutical forms.

A novel support for laccase immobilization: cellulose acetate modified with ionic liquid and application in biosensor for methyldopa detection.

A material based on cellulose acetate (CA) and the room temperature ionic liquid 1-butyl-3-methylimidazolium bis(trifluoromethylsulfonyl)imide (BMI·N(Tf)(2)) was developed and characterized by scanning electron microscopy, electron dispersive spectroscopy and infrared analysis. Laccase (Lac) from Aspergillus oryzae was immobilized in this material to investigate the behavior of methyldopa by square-wave voltammetry. Under optimized conditions, the Lac biosensor based on CA/BMI·N(Tf)(2) exhibited an excellent electrocatalytic performance: the analytical curve showed good linear range for methyldopa concentrations from 34.8 to 370.3 µM with a detection limit of 5.5 µM. This sensor demonstrated acceptable stability (ca. 60 days; at least 350 determinations), good repeatability and reproducibility (relative standard deviations of 1.5 and 4.3%, respectively). The recovery study of methyldopa in pharmaceutical formulations ranged from 94.1 to 105.9%. The determination of this substance using the biosensor compared favorably with that using a spectrophotometry procedure at the 95% confidence level, and indicated potential application to methyldopa determination in pharmaceutical samples.

Quantitative analysis of levodopa, carbidopa and methyldopa in human plasma samples using HPLC-DAD combined with second-order calibration based on alternating trilinear decomposition algorithm.

An HPLC method combined with second-order calibration based on alternating trilinear decomposition (ATLD) algorithm has been developed for the quantitative analysis of levodopa (LVD), carbidopa (CBD) and methyldopa (MTD) in human plasma samples. Prior to the analysis of the analytes by ATLD algorithm, three time regions of chromatograms were selected purposely for each analyte to avoid serious collinearity. Although the spectra of these analytes were similar and interferents coeluted with the analytes studied in biological samples, good recoveries of the analytes could be obtained with HPLC-DAD coupled with second-order calibration based on ATLD algorithm, additional benefits are decreasing times of analysis and less solvent consumption. The average recoveries achieved from ATLD with the factor number of 3 (N=3) were 100.1+/-2.1, 96.8+/-1.7 and 104.2+/-2.6% for LVD, CBD and MTD, respectively. In addition, elliptical joint confidence region (EJCR) tests as well as figures of merit (FOM) were employed to evaluate the accuracy of the method.

Enantioseparation of dopa and related compounds by cyclodextrin-modified microemulsion electrokinetic chromatography.

A chiral microemulsion electrokinetic chromatography method has been developed for the enantiomeric separation of 3,4-dihydroxyphenylalanine (dopa), its precursors phenylalanine and tyrosine, and the structurally related substance methyldopa. The separations were achieved using an oil-in-water microemulsion, which consisted of the oil-compound ethyl acetate, the surfactant sodium dodecylsulfate (SDS), the co-surfactant 1-butanol, the organic modifier propan-2-ol and 20mM phosphate buffer pH 2.5 or 2.0 as aqueous phase. For enantioseparation sulfated beta-cyclodextrin was added. The resolution of each racemate was optimized by varying the concentration of the buffer and all components of the microemulsion. Enantioseparation could be achieved for dl-dopa, dl-phenylalanine and dl-tyrosine within 13 min with a resolution of 4.3, 3.1 and 3.3, respectively, and for methyldopa in 17 min (Rs: 1.4). The established methods allowed the detection of dopa, phenylalanine, tyrosine and methyldopa with a limit at 0.5, 1.0, 0.2 and 2.0 microg/ml.

Simultaneous kinetic-spectrophotometric determination of carbidopa, levodopa and methyldopa in the presence of citrate with the aid of multivariate calibration and artificial neural networks.

The kinetic methodology based on the difference of reaction rates, is based on the reaction between a common oxidizing agents such as tris(1,10-phenanthroline) and iron(III) complex (ferriin, [Fe (phen)3]3+) in the presence of citrate and spectrophotometrically, monitoring the changes of absorbance at the maximum wavelength of 511 nm. Experimental conditions such as pH, reagents and citrate concentrations were optimized, and the data obtained from the experiments were processed by several chemometric approaches, such as artificial neural network (ANN) and partial least squares (PLS). A set of synthetic mixtures of carbidopa (CD), levodopa (LD) and methyldopa (MD) was evaluated and the results obtained by the applications of these chemometric approaches were discussed and compared. It was found that the back propagation artificial neural network (BP-ANN) method afforded better precision relatively than those of radial basis function artificial neural networks (RBF-ANN) and PLS. The proposed method was also applied satisfactorily to the determination of carbidopa, levodopa and methyldopa in real samples.

Cyclic voltammetric study of alpha-methyldopa at carbon paste electrode.

The cyclic voltammetric behaviour of alpha-methyldopa at a silicon oil carbon paste electrode has been reported. This allowed the development of a quantitative method to determine alpha-methyldopa in LiCl, KCl, NaCl, HCl, H2SO4 and CH3COOH as supporting electrolytes. For qualitative characteristics, alpha-methyldopa showed an ECC mechanism in terms of electron transfer reaction (placing it in DPI zone) at carbon paste electrode. The values of transfer coefficients alpha and beta were determined. The larger DeltaEp values were obtained due to the use of unmodified carbon paste electrode (CPE) has decreased the rate of electron transfer at the surface of the test electrode. The first order rate constant values (ko) were within 0.10-7.78 +/- 0.1x10(-3) s(-1). Adsorption of analyte was also determined at CPE using repeated scan method.

Spectrophotometric investigations of the assay of physiologically active catecholamines in pharmaceutical formulations.

Two simple, sensitive, and accurate spectrophotometric methods are proposed for the determination of levodopa (LD), methyldopa (MD), dopamine hydrochloride (DP), and pyrocatechol (PC) in pure and pharmaceutical preparations. The methods are based on measurement of the absorbances of tris(o-phenanthroline)iron(II) (method A) and tris(bipyridyl)iron(II) (method B) obtained by the oxidation of the catecholamines by iron(III) in the presence of 1,10-phenanthroline and 2,2'-bipyridyl at 510 and 522 nm, respectively. The absorbances were found to increase linearly with increases in the concentrations of the catecholamines, results which were corroborated by the calculated correlation coefficients (0.9990-0.9996). Beer's law was valid over the concentration ranges of 0.04-0.6, 0.06-0.75, 0.06-0.65, and 0.05-0.70 microg/mL in method A and 0.02-1.0, 0.04-1.3, 0.05-1.0, and 0.06-1.1 microg/mL in method B for PC, MD, LD, and DP, respectively. The common excipients and additives did not interfere in their determinations. The proposed methods were successfully applied to the assay of LD, MD, and DP in various dosage forms. The results were validated by statistical analysis.

Detection of alpha-methyldopa on thin-layer plates using pi-acceptors in 1,4-dioxane.

pi-Acceptors (P-chloranil and chloranilic acid) in 1,4-dioxane were used to detect alpha-methyldopa on thin layer plates. Thin layer plates coated to a thickness of 0.25 mm with silica gel G60 were spotted with solution of alpha-methyldopa from drug formulations and pure alpha-methyldopa solution. The plates were developed in a solvent system of methanol: glacial acetic acid: water (14:5:3). The developed plates were dried and sprayed with pi-acceptors, and further were counter-sprayed with dimethylformamide (DMF). P-Chloranil gave an immediate blue green complex with alpha-methyldopa which intensified on counter-sprayed with DMF. alpha-methyldopa (reference sample and those from drug formulations) gave Rf of 0.85 and excipients from the drug formulations did not interfere with the result.

Excretion of antihypertensive medication into human breast milk: a systematic review.

OBJECTIVE: To establish which antihypertensive medications are safe for use while breastfeeding, by reviewing the available evidence. METHODS: Reports of studies examining the transfer of antihypertensive medications to breastmilk were identified from multiple MEDLINE and EMBASE searches, manual review of bibliographies of articles and textbooks on drug use during lactation. The reports were reviewed and the results were compiled. RESULTS: Prospective cohort studies and case reports constituted the only available evidence. Compilation of these results found that the milk to plasma (M/P) ratios varied widely across the beta-blocker family, the beta-blockers with low protein binding having the highest M/P ratios. The angiotensin-converting enzyme (ACE) inhibitors, methyldopa, and some calcium channel blockers had low M/P ratios. CONCLUSION: The available data to date indicate that ACE inhibitors, methyldopa, beta-blockers with high protein binding, and some calcium channel blockers all appear to be safe treatments of hypertension in a nursing mother. The data suggest that drugs to be avoided are beta-blockers with low protein binding. However, the available evidence is limited and further studies are needed to confirm these findings.

The use of a response surface methodology on HPLC analysis of methyldopa, amiloride and hydrochlorothiazide in tablets.

A multifactor optimisation technique is successfully applied to develop a new HPLC method in which methyldopa, hydrochlorothiazide and amiloride were analysed and determined on a C18 column with detection at 286 nm. The optimal conditions of HPLC separation were determined with the aid of the response surface diagram -- 'window diagram'. The effect of simultaneously varying the pH, proportion aqueous acetic acidum and methanol in the mobile phase were studied to optimise the separation. The mobile phase composition that provides an acceptable resolution methyldopa, hydrochlorothiazide and amiloride in a short elution time is water--methanol (75:25) and pH 3.60. The k' values for methyldopa, hydrochlorothiazide and amiloride after optimisation were 1.40, 2.50 and 5.33, respectively. Relative retention (alpha) for ratio hydrochlorothiazide/methyldopa and amiloride/hydrochlorothiazide were 1.767 and 2.159, respectively. Correlation coefficients of the calibration curves for all analytes were greater than 0.995 and the R.S.D. values for the slope and the intercept with respect to the linearity were less than 2%. A method is applied for the quantitative analysis of Alatan tablets (Lek-Ljubljana). The powdered tablets are extracted with methanol, containing caffeine as the internal standard and assayed by comparison of peak areas after liquid chromatography. The high recovery (for all analytes about 100%) and the low R.S.D. (<2%) confirm good precision and reproducibility of the chromatographic method.

Direct high-performance liquid chromatographic determination of the enantiomeric purity of levodopa and methyldopa: comparison with pharmacopoeial polarimetric methods.

Chiral high-performance liquid chromatography was employed for determination of the enantiomeric purity of levodopa and methyldopa. The determination of D-DOPA in levodopa was accomplished using a chiral ligand-exchange chromatograpy with an ordinary C18 column and a chiral mobile phase containing N,N-dimethyl-L-phenylalanine and Cu(II) acetate or by means of LC on a teicoplanin column in conjunction with ethanol-water (65:35, v/v). Both methods gave good performance, however, the latter was faster and more convenient and suitable for routine analyses. For the determination of D-methyldopa a LC method based on the use of a teicoplanin column in polar organic mode with methanol-acetic acid-triethylamine (1,000:0.05:0.05, v/v/v) mobile phase was developed. The precision, accuracy, linearity and selectivity were satisfactory. In comparison with pharmacopoeial polarimetric methods (according to the European Pharmacopoeia and the Pharmacopoea Bohemoslovaca), the LC methods proved to be much more sensitive giving detection limits 0.04% of D-DOPA and 0.3% of D-methyldopa.

Quantitative microdialysis for studying the in vivo L-DOPA kinetics in blood and skeletal muscle of the dog.

In this study the microdialysis technique, using alpha-methyldopa as internal standard (IS), is introduced for the in vivo determination of L-DOPA, dopamine (DA), and their metabolites dihydroxyphenylacetic acid (DOPAC) and 3-O-methyldopa (3-OMD) in blood plasma and skeletal muscle extracellular fluid (ECF), in anaesthetised beagle dogs, after i.v. administration of L-DOPA. In a first calibration experiment, the in vivo relative losses (RL) of the compounds and the IS were determined. These were lower in skeletal muscle than in blood plasma. K was defined as the ratio of the RL of the IS to the RL of the compound of interest and was shown to be constant for a certain compound within one tissue. However, except for DA, a significant difference was seen in K values between blood plasma and skeletal muscle. In a second step, the method was validated in blood plasma. The AUC0-->3 values for the non-protein bound L-DOPA did not differ significantly between the dialysis (141.3 +/- 16.0 nmol.h/ml) and traditional whole blood sampling (145.3 +/- 18.7 nmol.h/ml), confirming that microdialysis combined with accurate calibration is a reliable technique for studying the kinetics of drugs in vivo in different tissues.

Pharmacokinetics and presystemic gut metabolism of methyldopa in healthy human subjects.

This study examined the pharmacokinetics and metabolism of methyldopa after giving single 250-mg oral and intravenous doses to 16 healthy human volunteers. A 48-hour washout period was allowed between oral and intravenous treatments. Blood and urine samples were collected; methyldopa was assayed in blood and urine, and its metabolites (methyldopa sulfate, alpha-methyldopamine, and alpha-methyldopamine sulfate) were assayed in urine. Pharmacokinetic parameters were recorded as follows: half-life was 2.0 +/- 0.7 hours; total body and renal clearance were 268 +/- 72 and 107 +/- 35 mL/min, respectively; and volume of distribution at steady-state was 33 +/- 11 L. The absolute bioavailability of the drug was 42 +/- 16%. The measurable metabolites in urine after oral and intravenous administration accounted for 27% and 17% of the dose, respectively. Methyldopa sulfate was the most abundant metabolite recorded; its quantity was higher after oral than after intravenous administration, 20.1 +/- 5.7% versus 6.7 +/- 5.3% of the dose (P < .05), suggesting significant presystemic gut metabolism. First-pass gut metabolism for methyldopa was estimated to be 17.6 +/- 6.9% of the dose given.

High-performance liquid chromatography with electrochemical detection for the determination of levodopa, catecholamines and their metabolites in rat brain dialysates.

A high-performance liquid chromatographic assay with electrochemical detection is described for the simultaneous determination of levodopa, 3-O-methyldopa, dopamine, dihydroxyphenylacetic acid, homovanillic acid, 3-methoxytyramine, noradrenaline, adrenaline, 3-methoxy-4-hydroxyphenylethylene glycol and 5-hydroxyindoleacetic acid in rat brain dialysates. Samples are obtained in vivo using the microdialysis technique. Microdialysis probes are placed in the brain area to be studied and neurochemicals are collected by perfusion of the probe with modified Ringer's solution. Direct injection of the dialysates allows rapid and reliable results to be obtained.

Stability-indicating high-performance liquid chromatographic assay for alpha-methyldopa in sustained-release capsules.

A stability-indicating high-performance liquid chromatographic assay has been developed for the analysis of alpha-methyldopa (MD) in sustained-release capsules and in the presence of MD decomposition products and an MD industrial impurity, 3-O-methyl-methyldopa (MMD). The method utilizes reversed-phase chromatography (cyano-bonded column), an acidic mobile phase containing sodium heptanesulphonate as ion-pairing reagent and UV detection. Detector responses were linear in the ranges 0.5-200 microgram/ml for MD and 0.2-100 microgram/ml for MMD. The mean recoveries of MD from authentic sample and sustained-release capsules were 100.09 +/- 0.38 and 100.38 +/- 0.46%, respectively. The recovery of MD added to degraded MD, were 99.69% by the proposed method and 153.13% by the US Pharmacopeial (USP) spectrophotometric method. The method is sensitive, accurate and rapid and can be used in routine analysis for MD.